The objective of this research is to understand how the mammalian cell regulates the concentration of the proteins that are externally oriented on the plasma membrane. Toward this objective, we have labeled by external labeling methods those proteins that constitute the bulk of plasma membrane proteins in hepatoma tissue culture (HTC) cells. We have analyzed by two-dimensional polyacrylamide gel electrophoretic methods the externally oriented proteins and glycoproteins of the plasma membrane. A complex set of glycoproteins is present on the plasma membrane, but these glycoproteins are not unique to this organelle. By cell fractionation methods and by enzyme probe analyses (with trypsin or neuraminidase) of intact and disrupted cells, we can show the existence of a significant intracellular membrane-bound reservoir of these same plasma membrane glycoproteins. The function of this intracellular reservoir of plasma membrane proteins in HTC cells forms a significant portion of the present research. These membrane glycoproteins are being isolated and purified in order to prepare specific antibodies. These antibodies are being used to study the route of biogenesis of these membrane proteins in cells not only growing normally in tissue culture but also after perturbation of the surface concentration of specific membrane glycoproteins by specific antibody or by hormones such as dexamethasone. The exchange of plasma membrane proteins with the proteins of the intracellular pool (via recycling) is also being analyzed. Homologous proteins are being identified on hepatocytes and are being assigned to their domain of residence on the hepatocyte plasma membrane. The turnover behavior of these proteins in the unique domains is being analyzed.